Methods of treating helminth infections with thiourea derivatives

ABSTRACT

1,1,3-TRISUBSTITUTED THIOUREAS, ACTIVE AGAINST HELMINTHS IN WARM-BLOODED ANIMALS, AND METHODS FOR THE PREPARATION AND USE THEREOF.

United States Patent 3,659,012 METHODS OF TREATING HELMINTH INFEC- TIONSWITH THIOUREA DERIVATIVES Herschel D. Porter, Lawrence Township, MarionCounty,

and Harold M. Taylor, Washington Township, Marion County, Ind.,assignors to Eli Lilly and Company,

Indianapolis, Ind.

No Drawing. Filed May 26, 1969,'Ser. No. 827,965

Int. Cl. A61k 27/00 US. Cl. 424-322 6 Claims ABSTRACT THE DISCLOSURE1,1,3-trisubstituted thioureas, active against'helminths in warm-bloodedanimals, and methods for the preparation and use thereof.

BACKGROUND OF THE INVENTION Heretofore, a number of compounds have beenused to combat helminth infections in warm-blooded animals. Among suchcompounds are the arylthioureas described by Shimotani, J. Pharm. Soc.Japan, 72, 328-330 (1952), and the thiocarbanilides described in SwissPat. No. 431,- 491 (1967). US. Pat. No. 3,395,233 (1968) describes aseries of phenylthiourea compounds effective against insects andacarids; however, these phenylthioureas showed no anthelmintic activitywhen tested against worms in warm-blooded animals. None of the prior artcompounds is effective against all species of helminths infesting warmblooded animals. Therefore, the search for more eifective anthelminticsis a continuing one.

SUMMARY A new series of therapeutic compositions has now been discoveredthat is efiectiveagain st a broad spectrum of helminth infections. Thenew anthelmintic compositions comprise a series of 1,1,34risubstitutedthioureas having the structure wherein R is naphthyl, phenyl, or asubstituted phenyl group R and R, when taken separately, represent C -Calkyl groups branched at the carbon attached to the nitrogen, or a C -CC- cycloalkyl group and can be the same or different;

R and R" when taken together with the nitrogen atom to which they areattached form a 5-, 6-, or 7-membered ring having a C -C alkylsubstituent attached to each of the carbon atoms in the ring adjacent tothe nitrogen, with or without additional C -C alkyl substituentsattached to other carbon atoms in the ring, such alkyl substituentsbeing the same or different.

3,659,012 Patented Apr. 25, 1972 DESCRIPTION OF THE PREFERREDEMBODIMENTS This invention is concerned with therapeutic anthelminticcompositions and processes. More particularly, this invention relates totherapeutic compositions containing one or more of the compoundsrepresented by the following structure:

wherein R is naphthyl, phenyl, or phenyl substituted with at least onesubstituent selected from the group consisting of C -C alkyl, C C,alkenyl, C 43 alkynyl, C C, aldroxy, C C alkylenedioxy, phenyl, phenoxy,hydroxy, amino, C -C monoalkylamino, C -C dialkylamino, C -C acyl,carboxy, C -C alkoxycarbonyl, carbamyl, C C; alkoxycarbonylamino, C -Cacyloxy, C -C a1- kylsulfonyl, sulfo, sulfamoyl, N-C1-c4 alkylsulfamoyl,N,N-C C dialkylsulfamoyl, halo, halomethyl, dihalomethyl, trihalomethyl,nitro and cyano;

R and R", when taken separately, represent C -C alky groups branched atthe carbon attached to the nitrogen, or a C -C C cycloalkyl group, andcan be the same or difierent;

R and R", when taken together with the nitrogen atom to which they areattached, represent a moiety ,of the formula N OHY).

o .X/ \H wherein n is a number from 2 to 4; X is a methyl, ethyl,napropyl, or isopropyl; and Y is hydrogen or C -C alkyl.

In the above formula, R represents a naphthyl, phenyl or phenylsubstituted group such as C -C alkylphenyl, C -C dialkylphenyl, C -Calkenylphenyl, C -C dialkenylphenyl, C -C alkynylphenyl, C -Cdialkynylphenyl, C -C alkoxyphenyl, C -C dialkoxyphenyl, C -Calkylenedioxyphenyl, biph'enyl, phenoxyphenyl, hydroxyphenyl,amino'phenyl, diaminophenyl, C -C3-(a-chloro-o-tolyl)-1,1-(1,6-diethyl-3 -methylhexamethylene -2-thiourea3- :,oz-dii0dO-P-IIOIY1) 1, 1-(1-ethyl-3-isopropyl-6-methylhexamethylene-2-thiourea l-cyclopentyl-1-isopropyl-3-(a,a, x-trifluoro-m-tolyl)2-thiourea 3- 2-chloro-p-tolyl) -1-cyclohexyl-1-( 1-methylpentyl)-Z-thiourea l-tert-butyl-1-cycloheptyl-3-(2,6-dichloro-p-tolyl)2-thiourea 1-sec-butyl-3- 2-chloro-3 ,S-xylyl) -1-(l-ethyll-methylpropyl -2-thiourea3-(4-sec-butyl-2-chlorophenyl)-1-(1-ethylbutyl)-1-(1- methylbutyl-2-thiourea 1-( 1,2-dimethylpropyl) -3- 3eth'yl-5-fiuorophenyl 1-1,1,2-trimethylpropyl)-2-thiourea 3- (2,6-difluoro-4-propylphenyl) -1-(l-ethyll-methylpropyl)-1-( l-ethylpropyl) -2-thiourea 1-(1,2-dimethylbutyl)-1- l-ethyl-l-methylpropyl -3-(4-isopropyl-2-nitrophenyl -2-thiourea 1-(1,1-dimethylbutyl)-3-(a:fluoro-2,4-xylyl)-l-tertpentyl-Z-thiourea 3- (a -chloro-a-fiuoro-2,5-xylyl) -l-cyclopentyl-1-cycloheptyl-Z-thiourea 3-(u ,a-diiodo-2,6-xylyl)-1,l-diisopropyl-Z-thioure-a 1,1-di-sec-butyl-3-(a ,a-difluoro-2,4-Xylyl)-2-thiourea 3- (a ,a -dichloro-a -fluoro-3,5-xylyl)-1,1-bis( l-methylbutyl) -2-thiourea 3- (a -chloro-other,a-trifluoro-2,4-xylyl) 1,1-dicyclohexyl-Z-thiourea1,1-(1,S-dimethylpentamethylene)-3-(a ,a ,a ,a ,a

pentachloro -3 ,4-xylyl -2-thiourea 1, 1- l,6-dimethylhexamethylene -3-a ,ot ,ot ,a ,ot ,a

hexabromo-2,6-x-ylyl -2-thiourea 3- (3-cyanophenyl)-1,1-di-tert-pentyl-Z-thiourea 3- (4-chloro-3-cyanophenyl) -1,1-bis(1,2-dimethylpropyl) -2-thiourea 3- (Z-cyano-p-tolyl)1,1-bis(l-ethylbutyl) -2-thiourea It is of the essence in the present inventionthat when R and R" are independent substituents that each carbon atomattached to the l-nitrogen must be branched, or part of a C C or Ccycloalkyl ring, and that when R and R" together with the nitrogen atomto which they are attached form a ring, each carbon atom attached to thenitrogen must bear a C -C alkyl substituent.

The compounds useful in this invention are generally crystalline solidmaterials which are generally soluble in many common organic solventsand relatively insoluble in water. The compounds exhibit the beneficialand desirable characteristics of high helminth toxicity and lowmammalian toxicity and are therefore very useful in treatinghelminthiasis in warm-blooded animals.

The compounds useful in this invention are prepared readily by allowingan isothiocyanate compound of the formula to react with an alkyl orheterocyclic secondary amine of the formula wherein R, R, R" are definedas above.

The reaction, common in the art, is conveniently carried out in thepresence of a suitable liquid reaction medium which is preferably anorganic liquid. Alternatively, when the amine reactant is a liquid atthe reaction temperature, it can be employed both as the reactant andthe liquid reaction medium. The reaction proceeds readily attemperatures between 0 C. and C., and preferably at temperatures between25 C. and 40 C., with evolution of heat generated from the exothermicnature of the reaction.

Isothiocyanate compounds appropriate for use in making the compoundsuseful in this invention can be prepared by reacting an aniline havingthe desired substituents attached thereto with an equimolecular amountof thiophosgene in a suitable reaction medium, such as 0.5 Nhydrochloric acid. The resultant isothiocyanate reaction product ispartitioned into diethyl ether and recovered therefrom by means known tothose skilled in the art.

Suitable organic liquids which can be used as reaction media includehydrocarbons such as benzene, toluene, xylene, and cyclohexane; etherssuch as diethyl ether and diisopropyl ether; alcohols such as methanol,ethanol, and isopropanol, chlorinated hydrocarbons such as chloroformand trichloroethylene; tertiary amines such as triethylamine andN-methylpiperidine; as well as solvents such as dimethyl sulfoxide,dimethylformamide, and the like.

The reactants can be employed in any amount, some of the desired productbeing obtained when the reactants are brought together in any ratio.Equimolecular amounts of the isothiocyanate and the secondary amine areconsumed in the reaction, and the use of amounts which represent suchproportions are preferred. When the secondary amine compound is employedas the reaction medium simultaneously with its use as a reactant, it ispreferable to use an excess of the reactant compound to maintain theliquidity of the reaction mixture.

The reaction is carried out by dissolving the isothiocyanate compound inone of the above-described organic solvents and then introducing thesecondary amine compound into the solution of the isothiocyanatecompound, with stirring, at such a rate as to control the rate ofreaction and to keep the temperature of the reaction mixture below 80 C.While some of the desired product is formed as soon as the reactants aremixed, the yield of the desired product is increased by stirring thereaction mixture for a period of time, preferably from about 30 minutesto about 2 hours. When the reaction is complete, the reaction productmixture is cooled to 0 C. for from 1 to 12 hours, with continuedstirring, to effect crystallization, after which the solid materialwhich separates is filtered off. The crude material is purified byrecrystallization. Solvents suitable for recrystallization includediethyl ether, ethanol, acetonitrile, dimethyl sulfoXide, benzene,petroleum ether, and the like.

The synthesis of the compounds useful in this invention is furtherillustrated by the following preparations, which preparations are not tobe construed as limiting the scope of this invention.

PREPARATION l 1,1-(1,S-dimethylpentamethylene)-3-(a,a,a-trifluorom-tolyl-2-thiourea About 10.2 gm. (0.05 mole) ofm-trifluoromethylphenyl-isothiocyanate were added to about 11.3 gm. (0.1mole) of 2,6-dimethylpiperidine, with stirring. The mixture became hot,and in about 10 minutes crystals began to appear. The reaction productmixture was allowed to stand at room temperature for about 30 minutes,cooled to 0 C. for 12 hours, and filtered. The filter cake was washedwith 10 ml. of cold commercial ethanol and then dissolved in 100 ml. ofhot commercial 95% ethanol containing 500 mg. of 2,6-dimethylpiperidine.The solution was cooled to C. for 12 hours, and the crystallineprecipitate filtered off and dried in vacuo at room temperature. About9.35 gmrof light yellow crystals melting at about 123-126 C. wereobtained and identified by elemental analyses and infrared spectrum as1,1-(1,5-dimethylepentamethylene) -3 (a,a,a-trifluoro-m-tolyl-2-thiourea 'The yield was 59.0 percent of theory.

Following the general procedure outlined in Preparation 1, and usingappropriate starting materials, the following compounds were prepared:

Product: Melting point, C.

1,1 (1,S-dimethylpentamethylene)-3-phenyl- Z-thiourea 78-80 3 4chlorophenyl)-1,1-(1,5-dimethylpentamethylene)-2-thiourea 120-124 1,1(1,5 dimethylpentamethylene)-3-(3- fluorophenyD-Z-thiourea 98-100 1,1(1,5 dimethylpentamethylene)-3-(3- nitrophenyl)-2-thiourea 135-136 3(2,5 dich1orophenyl)-1,1-(l,5-dimethylpentamethylene)-2-thiourea 87-89 3(3,4 dichlorophenyl)-1,1-(1,5-dimethylpentamethylene)-2-thiourea 138-1393 (2,4 dichlorophenyl)1,1-(1,5-dimethylpentamethylene)-2-thiourea125-128 3 (3 -'chlorop-tolyl)-l,l-(1,5-dimethylpentamethylene)-2-thiourea 122-124 1,1 1,5dimethylpentamethylene)-3-(ma ttrifiuoro-4-nitro-m-tolyl)-2-thiourea122-123 1,1 (1,5 dimethylpentamethylene)-3-(3,4- xylyl)-2-thiourea111-113 1,1- (1,5 dimethylpentamethylene)-3-(a,a,a-

trifiuoro-m-tolyl)-2-thiourea 124-128 3 (3 chloro ptolyl)-1,l-(1,5-dimethyltetramethylene)-2-thiourea 127-130 '1,1 (1,4dimethyltetramethylene)-3-(3,4-

xylyl )-2-thiourea 126-128 3 (6 chloro a,a,a-triflnoro-m-tolyl)-1,1-

( 1,S-dimethylpentamethylene) -2-thiourea 118-122 3 (41,11,111.trifluoro-m-tolyl)-l,l-(1,3,5-trimethylpentamethylene)-2-thiourea 80-83PREPARATION 2 1,1-di(sec-butyl)-3-(3-chloro-p-tolyl)-2-thiourea To asolution of 9.0 gm. (0.05 mole) of 3-chloro-4-methylphenylisothiocyanate in 100 ml. of diethyl ether were added, withstirring, 6.5 gm. (0.05 mole) of di-sec- 'butylamine. A mild exothermicreaction occurred. The reaction mixture was stirred for minutes, andthen 100 m1. of petroleum ether were added. The reaction mixture wascooled to room temperature, filtered, and the filtrate concentrated invacuo. The yellow oil thus obtained was dissolved in 100 ml. of hotpetroleum ether and the solution cooled to 0 C. for 24 hours. The solidprecipitate was filtered 011 and recrystallized from petroleum ether toyield elf-white crystals melting at about 57-59 C. and identified as 1,1di(sec butyl)-3-(3-chloro-p-tolyl)-2- thiourea. The yield was 38.0percent of theory.

Following the same procedure as in Preparation 2, and using suitablysubstituted isothiocyanates and secondary amines, the followingcompounds were prepared:

. Melting point, C.

2 3 (4 fluorophenyl) 1,1 diisopropyl 2- thiour'ea 108-110 3 (3cyanophenyl) 1,1 diisopropyl 2- thiourea 112-115 1,1 diisopropyl 3 (3nitrophenyl) 2- thiourea 126-131 1,1diisopropyl-B-(3-methylsulfonylphenyl)- Z-thiourea 134-137 1,1diisopropyl 3-(a,m,a-trifiuoro-o-tolyl)- 1 Z-thiourea 98-100 1,1diisopropyl 3 (a,a,a-trifiuoro-mtolyl)- Z-thiourea 84-86 1,1 diisopropyl3 (4 nitrophenyl) 2- thiourea 125-129 3- (3,4 dichlorophenyl)1,1-diisopropyl- Z-thiourea -118 3 (3 chloro p tolyl) 1,1 diisopropyl-2-thiourea 104-106 diisopropyl-Z-thiourea 78-81 1,1- diisopropyl3-(e,a,a-trifluoro-4-nitrom-tOIyD-Z-thiourea -130 1,1-diisopropyl-3-(3,4-xylyl)-2-thiourea 118-120 1 l-diisopropyl-3 l-naphthyl -2-thiourea 119-1 20 1,1-diisopropyl-3-(2-naphthyl)-2-thiourea 85-8 71,1-di(sec-butyl)-3-phenyl-2-thiourea 5 1-54 1,1 dicyclohexyl 3 (cacao:trifluoro-mtolyl)-Z-thiourea 108-111 1,1 dicyclohexyl 3 (4nitrophenyD-Z- thiourea -153 3 (3 chloro p-tolyl)-1,1-dicyclohexyl-2-thiourea g 90-93 1,1-dicyclohexyl-3- (3,4-xylyl) -2-thiourea 97-9 8 Ithas been found that the 1,1,3-trisubstituted thioureas described abovepossess outstanding anthelmintic activity when administered orally ineifective amounts to a parasitized host. The active compound can beadministered alone or in combination with other materials, such asmedicinal preparations, nutrients, inert substances, and the like. Thethiourea compound can be incorporated in a variety of pharmaceuticaldosage forms; for example, as a compressed tablet, filled capsule,granule, powder, liquid suspension, or the like. The solid dosage formsare particularly convenient to administer, and in a preferred embodimentof this invention, comprise compressed tablets or filled capsules inwhich the active compound is combined with compatible excipients such asstarch, lactose, microcrystalline cellulose, sodium lauryl sulfate,talc, magnesium stearate, sodium carboxymethylcellulo-se, sodiumalginate, and the like, in such amounts and pro portions as to provide aunit dose which can contain from about 5.0 mg. to 5000.0 mg. of theanthelmintic compound.

In another embodiment of this invention, the active compound can beincluded in a liquid suspension in which the vehicle can be comprised ofwater, an edible oil, or an edible polyol such as, for example,glycerine or 1,2- propanediol, to which suspension can be added any of avariety of additional excipients such as preservatives, suspendingagents, surfactants, flavoring agents, specific gravity-adjustingagents, and the like.

In still another embodiment of the present invention, the active1,1,3-trisubstituted thiourea compound can be administered orally in asuitable feed. Administering the anthelmintic compound in the feed offarm animals is preferably accomplished by preparing an appropriate feedpremix containing the thiourea and incorporating the premix into thecomplete ration. The premix can be prepared by mixing the activecompound with, for example, alfalfa grits, solvent-extracted soybeanfeed, corn meal, and the like, or a non-nutritive, nonirritatingmaterial which is acceptable tothe animal such as exfoliatedhydrobiotite, kaolin, talc, attapulgite, bentonite, microcrystallinecellulose, and the like, in high concentration. The premix is then addedto or mixed with the feed to provide a ration containing from about0.025 percent to about 2.5 percent of the anthelmintic compound.Alternatively, an intermediate concentrate or feed supplement containingthe thiourea compound can be blended into the feed.

The preferred method for adding the anthelmintic compound to the feed ofdomestic animals comprises the incorporation of from to 50 percent by'weight of the thiourea compound into a granule or powder in intimateadmixture with physiologically inert excipients, such as,illustratively, kaolin, bentonite, talc, sodium carboxymethylcellulose,starch, and the like, so that the proper unit dosage quantity in gramsmay be related to the conventional units of volume, such as tablespoon,cup, pint, and the like, readily understood by the average animal owner.Granules having a diameter of from 0.5 mm. to 2.0 mm. are preferredbecause of consistent density, flow characteristics, and ease of uniformblending throughout the feed in the desired amount.

The active thiourea compound can be given in a single dose or in divideddoses. When given as a single dose, the dosage of the active compoundcan range from about 5 to about 500 mg./kg. of body weight, preferablyfrom about 25 to about 100 mg./kg. A divided dosage regimen can comprisea total dose of from about 25 to about 2500 mg./ kg, or more, the exacttotal dose being a function of the amount in each treatment dose and theduration thereof. A method of choice for treating helminthiasis in dogsand cats comprises the administration of a single dose of, for example,100 mg./kg. ofl,l-(1,5-dimethylpentamethylene)-3-(a,u,a-trifluoro-m-tolyl)-2-thioureain the form of compressed tablets. A preferred method for treating farmanimals, such as poultry or swine, comprises adding an appropriateamount of a feed premix containing the active compound to the animalsdaily ration so as to provide for the ingestion by each animal of about100 mg./kg. of, for example 1,1-diisopropyl-3-(a,a,m-trifluoro-m-tolyl)-2-thiourea. Those skilled in the art will recognize that the actualdosage and mode of administration will be governed by the species, age,size, and general condition of the animal, the severity of the helminthinfection, and the convenience of the method of treatment.

The anthelmintic activity of the compounds useful in this invention hasbeen demonstrated in in-vivo tests.

against the dog large roundworms (ascarids), Toxocara cam's and Toxascaris leonina; dog hookworms, Ancylostoma caninum and Uncinariastenocephala; and the dog tapeworms, Taenia spp. and Dipylidium caninum.The thiourea compounds have also been shown by i-n-vivo tests in the ratto control the small roundworm species, Strongyloides ratti. Inaddition, the thiourea compounds useful in this invention have beenshown to possess invivo activity against the helminth speciesPhysaloptera spp. in dogs; Toxocara cati and Ancyclostomzz caninum incats; Ascaridia galli and H eterakis gallinarum in chickens;Trichostrongylus spp., Nematodirus spp., Moniezia spp., and Haemonchuscontortus in sheep; and Ascaris suum. in swine. In further tests, thelarval stage, microfilaria, of the dog heartworm, Dirofilaria immitis,circulating in the blood has been effectively controlled by theadministration of 25 mg./kg./day of1,1-(l,5-dimethylpentamethylene)-3-(a,a,a-trifluoro-m-tolyl)-2-thioureafor 29 days. While the mature worms are not killed, the larval stage ofthe infection is brought under control, thus eliminating a vector in thetransmission of the dog heartworm in the canine population.

Those skilled in the art generally recognize that agents which areeffective against species of helminths such as those listed immediatelyabove will also generally show activity against similar helminth specieswhich parasitize humans, such as the large roundworm (ascardis), Ascarislumbricoides; the hookwarm, necazor americanus; the tapeworm, Taeniasolium, Taenia saginata, and D-iphyllobothrium spp.; and the smallroundworm, Stronglyoides stercoralis.

The following examples further illustrate the present invention.

10 EXAMPLE 1 Evaluation of the anthelmintic properties of the 1,1,3-trisubstituted thioureas was carried out in vitro on the third-stageexsheathed sheep nematode, Haemonchutr contortus. The third-stage larvaewere suspended in buffered physiological saline of pH 7.2 in a dilutionthat provided about larvae per 0.1 ml. About 5.0 mg. of the testcompound were weighed into a 15-ml. test tube, to which was added onedrop of a 1:1 mixture of polyoxyethylene sorbitan monolaurate and water.About 0.45 ml. of acetone was added to the test tube to accomplishsolution of the test compound, and about 4.5 ml. of bufferedphysiological saline of pH 6.4 were added to complete the dilution toabout 1000 p.p.m. The testing at 1000 p.p.m. was carried out by placing0.9 ml. of the test compound solution and 0.1 ml. of the Haemonchuscontortus larvae suspension in a plastic well of a hemagglutinationplate and the medium incubated for 18 hours at 3738 C. When the test wasconducted at 100 p.p.m. of the test compound, 0.1 ml. of the testcompound solution, 0.8 ml. of buffered physiological saline, and 0.1 ml.of the Haemonchus contortus larvae suspension were added to the plasticwell and incubated as before. At the end of the incubation period, thetest in the plastic well was observed for worm motality.

The following listed compounds all decreased worm motility by at least70 percent at dilutions of 100 p.p.m. At the higher concentration, manyof the compounds completely suppressed worm motility.

COMPOUNDS 3-(2,4-dichlorophenyl) -1,1-( 1,5-dimethylpentamethylene-2-thiourea 3- (4-chlorophenyl)-1, 1-( 1,5 -dirnethylpentamethylene2-thiourea 1,1-( 1,5 -dimethylpentamethylene) -3- 3-fluorophenyl)2-thiourea 3- 2,5 -dichlorophenyl 1 1- 1,5 -dimethylpentamethylene-2-thiourea 3-(3,4-dichlorophenyl)-1,1-(1,5-dimethylpentamethylene-2-thiourea 1 1- 1,5 -dimethylpentamethylene) -3- 3,4-xylyl -2- thiourea1 3- 6-chloro-a,a,a-trifluoro-m-tolyl) l 1- 1,5-dimethylpentamethylene-2thiourea 1,1,-( 1,5 -dimethylpentamethylene -3- a,a,a-trifiuoro-4-nitro-m-tolyl -2-thiourea 1, l- 1,5 -dimethylpentamethylene -3-phenyl-2-thiourea 1,1-( 1,5 -dimethylpentamethylene -3-a,a',a-trifluoro-mtolyl -2-thiourea 1,1-( 1,5 -dimethylpentamethylene-3- a,ot,a-trifluoro-pto1yl)-2-thiourea3-('a,u,m-trifluoro-m-tolyl)-1,1-(1,3,5-trimethylpentamethylene)-2-thiourea 1,1-( 1,4-dimethyltetramethylene -3 a,u,a-trifiuoro-mtolyl-2-thiourea l, l-diisopropyl-3-phenyl-2-thiourea 1,1-diisopropyl-3-(l-naphthyl) -2-thiourea 1,1-diisoproply-3-(2-naphthyl)-2-thiourea 3-(4-acetylphenyl) -1,l-diisopropyl-Z-thiourea 1,1-diisopropy1-3-(Z-nitrophenyl)-2-thiourea' 3- (4-cyanophenyl)-1,l-diisopropyl-Z-thiourea 1,1-diisopropyl-3-(4-methoxyphenyl)-2-thiourea 1,1-diisopropyl-3 (3 -methoxyphenyl -2-thiourea1,1-diisopropyl-3-(p-tolyl)-2-thiourea 1,1-diisopropyl-3 cc,a,u-trifluoro-mtolyl -2-thiourea 1,1-diisopropyl-3-(m-tolyl)-2-thiourea1, l-diiospropyl- 3- (3 -methylsulfonylphenyl) -2-thiourea1,1-diisopropyl-3- (a,a,a-trifluoro-o-tolyl) -2-thiou'rea3-(6-chloro-a,a,a-trifiuoro-m-tolyl)-1,1,-diisopropyl-2- thiourea 1 1 1,l-diiso pro pyl-3- 3 ,4-xy1yl) -2-thiourea 3 -(3,4-dichloropheny1)-1,l-diisopropyl-2-thiourea 3-(2-fiuorophenyl)-l,1,-diisopropyl-2-thiourea3-(2,4,5-trichlorophenyl)-1,1-diisopropyl-2 thiourea 12 provided adlibitum. A single oral dose of 100 mgQ/kgl of body weight of thioureatest compound was administered to each dog, with the exception of thefirst two compounds listed in Table I, and marked with an asterisk. Each3-(3,5-dibromophenyl)-1,l-diisopropyl-Z-thiourea 5 of these twocompounds was administered at a dosage of1,l-diisopropyl-3-(3-nitrophenyl)-2-thiourea 350 mg./kg. of body weight.The dosage was prepared 1,1-diisopropyl-3-(4-nitrophenyl)-2-thiourea bygrinding the test compound to a fine powder in a'3-(3-cyanophenyl)-1,1-diisopropyl-2-thiourea mortar and pestle. Thefinely ground powder was filled3-(3-chloro-p-tolyl)-1,l-diisopropyl-Z-thiourea into large gelatincapsules which were administered oral-3-(4-fiuorophenyl)-l,l-diisoPropyl-Z-thiourea ly with or without aballing gun. Fecal material was col-3-(3-fluorophenyl)-1,l-diisopropyl-Z-thiourea lected daily for threedays after treatment and screened1,l-dicyclohexyl-3-(4-nitrophenyl)-2-thiourea for passed worms whichwere counted and identified3-(3-chloro-p-tolyl)-1,l-dicyclohexyl-Z-thiourea according to speciesand maturity. If nematode species1,l-dicyclohexyl-3-(3,4-xy1yl)-2-thiourea only were found in the feces,the dogs were sacrificed.1,1-dicyc1ohexyl-3-(a,m,ot-trifluoro-m-tolyl)-2-thiourea three daysafter treatment and any worms remaining in1,1-di(sec-butyl)-3-(4-nitrophenyl)-2-thiourea the gut recovered. Iftapeworms were identified in the1,l-di(sec-butyl)-3-(3-chloro-p-tolyl)-2-thiourea feces, the dogs weresacrificed 10 to '14 days after treatment. All of the worms remaining inthe gut of the EXAMPLE 2 animal at necropsy were then counted andidentified as to species and maturity. The number of Worms found at Y EP unwormed for at least 30 y Prior to necropsy was added to the numberof worms passed to the administration of the test compound, were used ingive the total count The percent f Worms removed by this critical test.The use of the word critical in conthe thiourea was computed by dividingthe number f nection with an anthelmintic test signifies that all of theeach helminth species passed by h total count, The Worms Passed in the69 were Ecol/cred, identified and common dog hookworm, Ancylostomacanirmm, was counted during the observation period and that the animalPresent in all th dogs sacrificed; and in addition, one or wassacrificed and necropsied and the number and species more f h followingspecies were found in the dogs: of worms remaining in the hostdetermined. Each dog Toxocara canis, Toxascaris leonina, Uncinaria stenoserved as its own standard. Each dog was placed in cephala, Trz'churisvulpis, Taenia spp., Dipylidiwm canan individual cage, allowed severaldays to adjust to his inum, and Physaloptera spp. Table I summarizes thereerwironment, and the presence of a natural helminth insults of thetreatments with representative 1,1,3 trisubfection confirmed byexamining the feces for parasitic stituted thioureas against the threemost frequently obova or tapeworm segments. Each dog was fed a dailyserved helminths, Toxocara canis, Ancyclostoma caninum, rationsuflicient for the size of the dog used, water being and Uncz'nariastenocephala.

TABLE I Anthelmintie efficacy oi representative 1,1,3 trisubstitutedthioureas in dogs 1 Percent worms removed, helminth No. dogs TozocaroAncylostoma Total Worms 8 M--' Mature; I'=Immature. -=No worms of thespecies were observed.

Compound I tested cams caninum ste g filz1,1-diisopropyl-3-pheuyl-2-thiomca 4 88 a 86 1003-(3,4-dichlorophenyl)-1,1-dilsopr0pyl-2-thiouroa. 4 100 M 10 1003-(3-fiuorophcnyl)-l,l-diis0propyl-2-thiouroa a 5 as 1001,1-diisopropyl-3-(a,m,a-trifluoro-m-tolyl)-2-thiourca 4 y 100 1003-(m,,a,a,aflaf-hexafiuoro-3,5-xylyl)-1,l-diisopropyl-Z-thiourea. 3 E885 0 3-(3 chloro-p-tolyl)-1,l-diisopropyl-Z-thiourea-" 2 {B 100 I 1003-(3 cyranophenyl)-1,1-diisopropyl-2-thiourea 2 951,l-dfisopropyl-ii-t3-methoxypheny1)-2-thiourea 2 I? 52 103(ti-chlordu,d,a-trifiuoro-m t01yl)-1,1-diisopropyl-2-thiourca 2 g: 1003-(2-fluorophenyl)1,I-diisopropyl-Mhiourea .Q. 2 figg 761,l-diisopropyl-d-(a,a,a-trifluoro-o-tolyl)-2-thiourea- 2 at so 2r(3,5dibromophenyl)-1,1-diisop1opy1-2-thiourca 1 V 1001,1-(1,fi-dimethylpentamothylenc)-3-(a,a,a-triflu0ro-1u-toIyl)-2-thioureas i 99+ 100 1,1-(1,fi-tlimethylpentamethyleno)-3-pl1enyl-2-thiourea -l 267 89 3-(2,5-dieh1orophenyl)-1,1-(1,5 dimethylpentamethylene)-2-thiourea.Q 2 35 100 B-(G-ehlom,a,atrifluorom-to1yl)-1,1-(1,o-dimcthylpentamethylene)-2-tl1i0urea 13-(4-chlorophenyl)-1,1-(1,-dimethylpentamethylene)-2-thiourea 1 100 1003-(3,4-dichlorophenyl)-1,1-(1,5-dimethylpentamethylene) 2-thiourea 4 i l78 100 3-(a,a,-trifluoro-mtolyl)-1,1-(1-,3,li-trimethylpentamethylene)'-2-thi0urea 1 510o 10o 1,1'diisopropyl-li (a,a,a-trifluoro-m-tolyl)-2-thiourea..t -lr.8 i 94 1,1-dl(sec-butyll-It-(dehloro-p-tolyl)-2-thiourea. 1 {T fi 25 13EXAMPLE 3 TABLE II Anthelmintie efiicacy of1,1-(1,54iimethylpentamethylene)-3-(u,a,atrifluoro-m-tolyl)-2-thioureain dogs 1 Percent wonns removedfl helmlnth No. dogs Torocam AncylostomaUnciuaria Dosage form tested cam's ca'nmum stenocephala Filled capsules3 99 i; 97 91 Compressed 67 M tablets. 19 {21 I i 96 100 1 Single oraldose of 100 mgJkg. of body weight.

Worms Passed -X100.

Total Worms 3 Number includes the 6 dogs reported in Table I.

4 M=Matnre; I=Immature.

2 Percent Worms Removed EXAMPLE 4 The compound 1,1 (1,5dimethylpentamethylene)-3 (a,a,a-trifiuoro-m-tolyl) -2-thiourea wasadministered orally to two dogs over a period of several days as shownin Table III. Both dogs were found to be infected with the heartworm,Dirofilaria immitis, by a quantitative examination of the blood formicrofilaria, the larvae stage of the heartworm. Each dog was treatedonce each day with an oral capsule. The absorption of therapeuticallyeffective amounts of the test compound into the blood stream of theanimals is shown by the reduction in the number of microfilaria in theblood over the duration of the treatment. The mature heartworms are tobe found in the heart chambers of the animal, while the larvae of theworm inhabits the blood. It is suggested by the data that dailyadministration of the test compound for several days is advantageous inachieving efficacy against the microfilaria.

TABLE III Efficacy of 1,1(1,S-dimethylpentamethylene)-3-(a,a,u-trifluoro-m-tolyl)- 2-2-tliioreauagainst the microfilaria of the dog lieartwonn larvae within livefemales.

1 Larval stage of heartworm. 3 Mature lieartworm.

EXAMPLE 5 A medicated feed suitable for chickens is prepared byincorporating the test compound, l,l-diisopropyl-3-(u,u,atrifluoro-m-tolyl)-2-thiourea, in a feed premix andblending the thus prepared premix with a ton of ration to provide achicken feed containing 2000 gm. of the active compound per ton.

The premix is prepared by grinding the active compound to a fine powderand mixing 100 gm. of the thus prepared compound with 344 gm. ofsolvent-extracted soybean meal, adding thereto gm. of mineral oil tostick the ingredients together and reduce the dustiness of the premix.

The premix prepared as illustrated above is intimately blended with thecomponents of a chicken ration to pro- 14 duce a feed containing 2000gm. active compound per ton according to the following formula:

Ingredient: Quantity, lbs. Corn, yellow, ground 1142 Soybean oil meal,solvent extracted (50%) 360 Corn gluten meal (60%) 100 Distillers driedsolubles, corn 100 Fish meal with solubles 100 Meat scraps (55%) Alfalfameal, dehydrated (17%) 38 Animal fat, beef tallow 20 Dicalciumphosphate, feed grade 12 Calcium carbonate (ground limestone) 10 Salt(NaCl) 6 Anthelmintic premix 20 Trace mineral premix 1 2 Vitamin premix2 10 Total 2000 Each pound contains: manganese, 30.4 mg; zinc, 34.0 mg.;Iron 7.7 mg.; copper, 0.8 mg.; and iodine, 0.4 mg.

Each pound contains: 450,000 I.U. Vitamin A; 120,000 I.C.U. Vitamin D3;1000 LU. Vitamin E; Menadione Sodium Bisulfate, mg.; Riboflavin, 400 mg.Niacin, 3.600 mg.; d- Pantothenic Acid, 966 mg.; Choline, 26,037 mg.;Vitamin Bu, 1 mg. Antioxidant, 2.5%.

EXAMPLE 6 A medicated granule suitable for domestic pets is prepared asfollows: 10 kg. ofl,1-(1,5-dimethylpentamethylene)-3-(u,a,a-trifiuoro-m-tolyl)-2-thiourea,75 kg. talc, 10 kg. bentonite, 2.5 kg. sodium carboxymethylcellulose,and 2.5 kg. of starch are intimately blended in an appropriate mixer,followed by the addition thereto of about 20 liters of water withthorough mixing.

The mixture thus prepared is extruded into long cylindrical strands,having a diameter from about 0.5 to about 2.0 mm., by forcing themixture by mechanical pressure through a plate or die in which one ormore holes having a diameter of from about 0.5 to about 2.0 mm. areprovided. Machinery for this purpose is conventional used in theplastics extrusion trade, is readily available, and known to thoseskilled in the art. The moist strands, or extrudate, are introduced intoa machine of the type designed by Nakahara and described in US. Pat. No.3,277,520, Method and Apparatus for Making Spherical Granules, andidentified as a Marumerizer (a trademark of Fuji Denki Kogyo, Osaka,Japan), and converted into spherical granules of a diameter from about0.5 to about 2.0 mm. The spherical granules so formed are dried byconventional means to remove excess water. Desirably, about 10 to 20percent by weight of the spherical granules are included in either dryor wet dog or cat food at the time of each feeding of the animal. Thepet consuming such medicated food in normal amounts receives a dose ofabout 75 to about mg./kg. of body weight.

EXAMPLE 7 The preparation of 1000 filled capsules containing 250 mg. of1,1-(1,S-dimethylpentamethylene) 3 (mega-litifiuoro-m-tolyl)-2-thioureaper capsule is carried out by blending 250 gm. of the active compound,300 gm. of starch powder, and 10 ml. of mineral oil and filling suchmixture into No. 0 capsules in the amount of 560 mg. per capsule.

EXAMPLE 8 Compressed tablets containing 90 mg. of 1,1-(1,5-dimethylpentamethylene) 3 (u,a,a-trifluoro-m-tolyl)-2- thiourea per tabletsuitable for oral administration are made by the following procedure: 90gm. of l,1-(1,5-dimethylpentamethylene) 3 (a,ot,oc-tIlfiu0l0I11-tOlyl)-2thiourea are mixed with 484 gm. of lactose, 6.4 gm. of starch, and 0.3gm. of magnesium stearate, and the mixture is dry graulated by pressinginto large tablets weighing about 25 to about 40 gm. or more. The largetablets are reduced to granules of a size from about 8 to about 20 meshby processing through an oscillating granulator,

- known to'those skilled in the art. The resultant dry granules arepressed into tablets of size such that'each tablet contains 90 mg. ofthe active agent. The tablets are pref- EXAMPLE 9 A suspension of1,1-(1,5-dimethylpentamethylene)-3- (a,a,u-trifluoro-m-tolyl}-2-thioureacontaining 100 mg. of the active compound per ml. of suspension isprepared by the following procedure. About 500 ml. of purified water areplaced in a suitable container and 5 gm. of sodiumcarboxymethylcellulose are hydrated therein, following which 1.0 gm. ofsoluble saccharin, 10.0 gm. of sodium cyclamate, 10.0 gm. ofsodiumcitrate, 0.75 gm. of methyl-p hydroxybenzoate, and 250 gm. of granulatedcane sugar are dissolved therein. To this mixture are added -0.1 ml. ofoil of anise and 250 mg. of vanillin, with stirwherein R is naphthyl,phenyl, or phenyl'substituted with at least one substituent selectedfrom the group consisting of C -C alkyl, C -C alkenyl, C -C alkynyl, C Calkoxy, C -C alkylenedioxy, phenyl, phenoxy, hydroxy, amino, C C,monoalkylamino, C -C dialkylamino, C -C alkanoyl, carboxy, C -Calkoxycarbonyl, carbamyl, C C, alkoxycarbonylamino, C -C alkanoyloxy, C-C alkylsulfonyl, sulfo, sulfamoyl, N-C -C alkylsulfamoyl, N,N-C -Cdialkylsulfamoyl, halo, halomethyl, dihalomethyl, trihalomethyl, nitro,and cyano;

R and R", when taken separately, each independently represents a memberselected from the group consisting of C -C alkyl branched at the carbonattached to the nitrogen, cyclopentyl, cyclohexyl, and cycloheptyl.

2. The methodof claim 1 wherein the compound administered is1,1-diisopropyl-3- (u,oc,u-t1'iflllOI0m-t01y1)2- thiourea.

, 3. The method of claim 1 wherein the compound administered is1,l-dicyclohexyl-3-(a,a,a-trifluoro-m-tolyl)- 2-thiourea. I

4. The method of claim 1 wherein the compound administered is 1-,1di(sec butyl)-3-(S-chloro-p-tolyD-Z thiourea.

5. The method of claim 1 wherein the compound administered is1,1-diisopropyl-3-(3,4-xy1yl)-2-thiourea.

6. The method of claim 1 wherein the compound administered is3-(3,4-dichlorophenyl)-1,l-diisopropyl 2- thiourea. I

References Cited UNITED STATES PATENTS 3,266,982 8/ 1966 Popofit 424-3223,365,360 1/1968 Taylor 424322 3,376,194 4/ 1968 Berger et al. 424-322SAM ROSEN, Primary Examiner US. Cl. X.R.

